SPRY1基因与角质形成细胞衰老的相关性研究

Significance of SPRY1 gene in senescence of keratinocytes

目的:探讨软脂酰化磷蛋白Sprouty1(SPRY1)基因在不同年龄段人体皮肤角质形成细胞和人永生化角质形成细胞(HaCaT细胞)衰老中的表达及意义。方法:收集不同年龄段正常皮肤标本120例,分为少年儿童组(<18岁)、青年组(18~44岁)、中年组(45~59岁)、老年组(≥60岁),各30例。通过免疫组化、Real-time PCR、Western blot技术检测SPRY1在各年龄段皮肤角质形成细胞中的定位及表达差异。用200μmol/L H2O2处理HaCaT细胞构建细胞老化组模型。Real-time PCR技术和Western blot技术检测SPRY1在正常组和衰老组HaCaT细胞中的表达。结果:SPRY1的表达主要分布在皮肤角质形成细胞的棘层及颗粒层,呈细胞质分布;SPRY1 mRNA及蛋白在人皮肤角质形成细胞中的表达随着年龄段的增长呈逐渐增高的趋势,差异具有统计学意义(P<0.05)。HaCaT细胞经H2O2处理后发生了老化。SPRY1 m RNA及蛋白在老化后的HaCaT细胞中表达上调,差异具有统计学意义(P<0.05)。结论:SPRY1基因在人体衰老的皮肤角质形成细胞及老化HaCaT细胞中表达增加,提示SPRY1基因参与角质形成细胞衰老调控。

Objective:To investigate the expression and significance of SPRY1 gene in the senescence of normal human epidermal keratinocytes(NHEKs)and immortalized keratinocytes(HaCaT cells)from individuals of different ages.Methods:A total of 120 normal skin specimens were collected from individuals of different ages and divided into juvenile group(<18 years),youth group(18-44 years),middle-aged group(45-59 years old),and elderly group(≥60 years),with 30 specimens in each group.Immunohistochemistry,real-time PCR,and Western blot were used to compare the localization and expression of SPRY1 in keratinocytes between different age groups.HaCaT cells were treated with 200μmol/L H2 O2 to establish a model of cellular senescence.Real-time PCR and Western blot were used to measure the expression of SPRY1 in the groups of normal and senescent HaCaT cells.Results:SPRY1 was mainly expressed in the spinous and granular layers of NHEKs and was distributed in the cytoplasm.The m RNA and protein expression of SPRY1 in keratinocytes gradually increased with the increase in age(P<0.05).Senescence of HaCaT cells was observed after H2 O2 treatment.The mRNA and protein expression of SPRY1 was significantly upregulated in senescent HaCaT cells(P<0.05).Conclusion:There is an increase in the expression of SPRY1 gene in senescent NHEKs and senescent HaCaT cells,suggesting that the SPRY1 gene is involved in the regulation of keratinocyte senescence.